PROJECT SUMMARY Fractalkine protein (FKN, CX3CL1) suppresses microglial activation leading to decreased neurodegeneration in a number of neurological disorders, including Alzheimer's disease (AD). In AD, a large reduction in FKN has been observed in postmortem cortical tissue. We have studied FKN and its role in neurodegeneration extensively, and we have demonstrated a beneficial effect of FKN in multiple neurodegenerative disease models. Our work demonstrates that overexpression of a soluble form of FKN (sFKN) decreases tau pathology in Tg4510 mice. More importantly, we observed that sFKN significantly reduces brain atrophy and neuron loss in these mice. In Parkinson's disease (PD) mouse models, overexpression of sFKN, but not a cleavage-resistant, insoluble mutant FKN, reduces disease pathology and neuron loss. Positive activity with soluble versus membrane-bound FKN suggests that a soluble molecule has the potential to activate the receptor and achieve neuroprotective effects. These observations prompted the hypothesis that the FKN pathway could be exploited as a therapeutic approach to treat AD. High-throughput screening produced a series of chemical scaffolds that are novel small molecule agonists of CX3CR1 suitable for further optimization through this funding opportunity. The lead compound is potent and highly selective, and has been shown to reduce LPS-mediated activation of microglia. Here, we propose to develop orally bioavailable potent CX3CR1 agonist compounds suitable for use as chemical probes. This campaign leverages the unique resources of Sanford Burnham Prebys and USF Health. Chemistry efforts will refine the current lead compound to improve potency and selectivity, and enhance the compound's drug-like properties. These efforts are supported by a robust testing funnel consisting of cellular and biochemical assays of CX3CR1 signaling, as well as appropriate counter-screens. The physicochemical and pharmacological properties of the compounds will be optimized, followed by pharmacokinetic studies in rodents. We will test for target engagement and efficacy in the AD animal models, in which we will identify at least one CX3CR1 agonist with potent efficacy in vivo.